Environmental factors, such as light of different spectral compositions and temperature, can change the level of activated photoreceptors which, in turn, can affect the biosynthesis of secondary metabolites in the cells of green fruit. By briefly irradiating the harvested fruit of Capsicum annuum L. hot peppers with red light (RL, maximum 660 nm) and far-red light (FRL, maximum 730 nm) and by keeping them at a low temperature, we attempted to determine whether the state of phytochromes in fruit affects the biosynthesis of secondary metabolites. Using HPLC, we analysed the qualitative composition and quantitative content of the main carotenoids and alkaloids and the chlorophylls and ascorbate, in pepper fruit exposed to the above factors. We measured the parameters characterising the primary photochemical processes of photosynthesis and the transcript levels of genes encoding capsaicin biosynthesis enzymes. The total carotenoids content in the fruit increased most noticeably after 24 h of RL irradiation (more than 3.5 times compared to the initial value), and the most significant change in the composition of carotenoids occurred when the fruit was irradiated with FRL for 72 h. The capsaicin alkaloid content increased markedly after 72 h of FRL irradiation (more than 8 times compared to the initial value). It was suggested that decrease in the activity of phytochromes due to a low temperature or FRL may result in an increase in the expression of the PAL and CAM genes.